This invention relates generally to genetic engineering, and more specifically, to methods of increasing infection efficiency.
Scientists now routinely introduce genetic material into prokaryotic or eukaryotic cells to obtain the expression of heterologous genes. The application of this technology to human cells forms the basis for the burgeoning field of gene therapy. Typical methods include incorporating DNA directly into cells, for example, by calcium phosphate precipitation, and using vector-mediated infection. Scientists have used a wide variety of viruses as gene-transfer vectors. In particular, non-competent retroviruses have proved useful because they allow the stable introduction of genetic material into the chromosome of the infected target cell.
The process of retroviral infection involves many steps. In the initial step the virus comes in contact with the cell. Next, the viral RNA is internalized. Reverse transcription follows. Then the viral DNA enters the nucleus. Finally the virally derived DNA is integrated into the target cell's chromosomes. This step is believed to require cell division.
Retroviruses are assembled by so-called packaging cell lines. Scientists have developed two methods to infect a target cell with a retrovirus. First, one may co-culture the target cell with the packaging cell line. This approach has proven successful, but unfortunately it represents the clinically less attractive procedure. There is the risk of contaminating the infected target cells with cells from the packaging line, with subsequent risks.
Second, one may culture a bed of target cells and add to them spent medium from the packaging cell line. Scientists believe that this approach is limited by the concentration of the virus in the spent medium. It has proven difficult to obtain high titer supernatants from packaging cell lines because the viruses tends to disintegrate rapidly. In any case, attempts to increase the concentration of the virus has not led yet to a dramatic increase in rates of infection.
Thus, there exists a need for methods to increase the rate of infection of target cells by retroviral vectors. The present invention satisfies this need and provides related advantages as well by providing methods to direct to motion of vectors towards the target cells.